Stress-sensitive microRNAs that play regulatory roles in bone formation
A stress, inhibitor-103a technique for biological applications
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Embodiment 1
[0186] Example 1. Loss of load leads to loss of bone mass in the hindlimb unloading mouse model Disuse osteoporosis
[0187] In order to study the influence of lack of mechanical stress stimulation on the homeostasis of bone remodeling, the inventors constructed a mouse model of unloading of both lower limbs (Hindlimb unloading, HU), which was originally developed by the National Aeronautics and Space Administration (National Aeronautics and Space Administration, NASA) It was built in 1980 to study the influence of weightlessness in space on the bone mass of astronauts, and then it was widely used in various studies of mechanics and motion systems ( figure 1 a). Compared with the mice in the baseline group (BS) and the static control group (Weight-bearing group, WB), the mice in the HU group were suspended by their tails for 28 days to suspend their hind limbs, but their forelimbs could walk freely without affecting their Drink water and eat. During the experiment, mice in e...
Embodiment 2
[0189] Example 2, CMS-mediated regulation of Runx2 post-transcriptional level in osteoblast differentiation
[0190] In order to explore the regulatory effect of CMS on osteoblast differentiation, the inventors used finite element analysis (FEA) to detect the load bearing conditions of the proximal femur ( figure 2 a). According to the existing literature reports, the stress carried by the normal human tissue level is usually less than 1000με. The FEA test found that the stress on the proximal femur was about 815±57με( figure 2 a). However, the load at the tissue level and the cell level are not the same concept. There is an amplification mechanism when the stress load is transmitted from the tissue level to the cell level, which is the stress-strain amplification mechanism at the cell level. In osteoblasts, the stress carried at the bone tissue level can be amplified 20 to 100 times when it is transmitted to the cell membrane. Therefore, the inventors selected 8% CMS (8...
Embodiment 3
[0192] Example 3. miR-103a directly targets Runx2 in osteoblast differentiation mediated by mechanical stress stimulation
[0193] Bioinformatics found that the 3'UTR of Runx2 is about 3777 nucleotides in length. The following miRNA bioinformatics prediction software: TargetScan, miRDB, miRanda and miRBase databases were selected to screen out miRNAs with potential targets in the 3'UTR region of the Runx2 gene, and their intersection was taken. Eliminating the miRNAs that have been reported to have a definite regulatory effect on Runx2, 12 miRNAs that can bind to the Runx2 3'UTR seed region (seed region) were obtained, including: miR-7, miR-22, miR-23b, miR-103a, miR-107, miR-143, miR-154, miR-221, miR-320d, miR-374b, miR-375and miR-384( image 3a).
[0194] In order to verify whether the screened 12 miRNAs are regulated by mechanical stress stimulation, the inventors loaded the hFOB1.19 human osteoblast cell line with 8% CMS for 3 days, extracted miRNAs, and detected the 12...
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